Chemokines and their receptors play a large role in mediating inflammatory responses to injury and disease. Monocyte chemoattractant protein-1 (MCP-1) is a chemokine that, through activation of its cognate receptor, the GPCR C—C chemokine receptor 2 (CCR2), has been implicated in the induction and maintenance of chronic pain. A six amino acid domain within the N-terminal domain of CCR2 binds MCP-1 with high affinity. Once bound, MCP-1 serves as a tethered ligand, and receptor signaling is mediated by secondary points of contact between the ligand and receptor, likely in the extracellular loop and transmembrane domain region of the receptor.
Toward an understanding of MCP-1 and CCR2 binding and signaling, Applicants synthesized and expressed a chimeric dog-human CCR2B molecule that has an overall 85% identity to human CCR2B. Positions 1-35 (N-terminal domain) and 301-360 (TM7 and C-terminal domain) of the chimeric receptor are equivalent to the human residues, while the intervening 265 residues are equivalent to the dog residues. Due to the relatedness between dog and human CCR2B, 211 of these 265 residues are identical between human and dog CCR2B, with the greatest number of mismatches mapping to EC2 and EC3.